Developments in the tools and methodologies of synthetic biology

Abstract

Synthetic biology is principally concerned with the rational design and engineering of biologically based parts, devices, or systems. However, biological systems are generally complex and unpredictable, and are therefore, intrinsically difficult to engineer. In order to address these fundamental challenges, synthetic biology is aiming to unify a "body of knowledge" from several foundational scientific fields, within the context of a set of engineering principles. This shift in perspective is enabling synthetic biologists to address complexity, such that robust biological systems can be designed, assembled, and tested as part of a biological design cycle. The design cycle takes a forward-design approach in which a biological system is specified, modeled, analyzed, assembled, and its functionality tested. At each stage of the design cycle, an expanding repertoire of tools is being developed. In this review, we highlight several of these tools in terms of their applications and benefits to the synthetic biology community.

Keywords: design cycle; engineering biology; standardization; synthetic biology; tools.

Figure 1

Synthetic Biology Index of Tools and Software (SynBITS). A schematic summary of the synthetic biology design cycle tools as depicted in SynBITS (www.synbits.co.uk), an online community-managed index of synthetic biology tools and software.

Figure 2

DNA assembly strategies. Restriction enzyme – restriction enzymes recognize specific DNA sequences and either cut within their recognition sequence (Type II) or adjacent to its recognition sequence (Type IIS) to create sticky or blunt-ended DNA fragments that can be ligated to other DNA fragments. Recombination – cellular DNA repair and recombination machinery can be utilized to integrate a DNA construct within a specific genomic locus. Integration is guided through 5′ and 3′ sequence complementarity of the integration sequence with the target locus. Overlap-directed – assembly order is guided by 20–40 bp overlaps at the ends of each DNA fragment that share sequence homology with adjacent DNA fragments. In the case of Gibson assembly, these homologous ends are processed (chew-back) and fused together (anneal) via the sequential activity of an exonuclease, a ligase, and a polymerase. DNA synthesis – DNA sequences are designed and optimized in silico for de novo synthesis. Commercial constructs are delivered as gene fragments or are pre-cloned within a plasmid vector.

Figure 3

Systematic design of biological systems. The biological design cycle is one of several engineering principles that have been adopted in synthetic biology, and it describes the iterative process of designing a biological system through multiple rounds of design, build, and testing. To ensure that iterations of the design cycle are informative, the systematic capture, and integration of experimental and experiential data within a biological design workflow, such as the one shown here, is desirable.